生物活性玻璃-丝素蛋白复合膜支持人牙髓干细胞增殖与分化初探  

Bioactive glass 45S5-silk fibroin membrane supports proliferation and differentiation of human dental pulp stem cells

作　　者：吕孝帅 李正茂[1] 王海燕[1] 杨雪超[1] 

机构地区：[1]广州医科大学附属口腔医院数字化口腔医疗中心·广州口腔病研究所·口腔医学重点实验室,510140

出　　处：《中华口腔医学杂志》2015年第12期725-730,共6页Chinese Journal of Stomatology

基　　金：广东省医学科研基金（A2014328）;广东省教育厅特色创新项目（2014KTSCX103）;广东省科技厅自筹经费类科技计划项目（2013-53）

摘　　要：目的 探讨生物活性玻璃45S5-丝素蛋白（bioactive glass 45S5-silk fibroin, BG45S5-SF）复合膜对人牙髓干细胞（human dental pulp stem cell,hDPSC）生长、增殖、分化的影响,为牙髓-牙本质复合体再生提供新思路和新方法.方法 制备纯丝素蛋白膜（蛋白膜组）、含不同质量浓度（1000和5000 mg/L）BG45S5-SF复合膜（复合膜A组和B组）,以无材料膜的孔板为空白对照组.接触角仪测量蛋白膜组和复合膜组表面接触角（每组3个复孔）.4组预孵24 h后接种hDPSC,培养第4、7、14、21天检测hDPSC增殖能力;扫描电镜和免疫荧光染色观察hDPSC黏附和生长;碱性磷酸酶活性评估hDPSC分化潜能;实时定量PCR检测hDPSC的成牙本质细胞向分化基因表达水平.结果 蛋白膜组、复合膜A组和B组表面接触角分别为89.51°±0.12°、70.32°±0.07°和71.31°±0.09°.hDPSC在丝素蛋白膜及BG45S5-SF复合膜上均黏附生长良好.复合膜A、B组第7、14、21天碱性磷酸酶活性及分化基因较空白对照组和蛋白膜组显著上调（P＜0.05）.复合膜A组牙本质基质蛋白1、牙本质涎蛋白、碱性磷酸酶、骨钙蛋白mRNA表达第14天达峰值,复合膜B组第21天达峰值;复合膜A、B组骨涎蛋白mRNA表达均在第21天达峰值.结论 BG45S5-SF复合膜不仅能支持hDPSC黏附、生长与增殖,还能促进hDPSC的成牙本质细胞向分化,提示BG45S5-SF复合膜有促进牙髓-牙本质复合体再生的潜能.Objective To investigate the effect of bioactivity glass 45S5-silk fibroin（BG45S5-SF） membrane on growth, proliferation and differentiation of human dental pulp stem cells（hDPSC）, and to provide new ideas and method for the regeneration of pulp-dentine complex.Methods hDPSC seed on pure silk fibroin membrane （protein membrane group） and BG45S5-SF membrane with different concentrations（1 000, 5 000 mg/L, composite membrane group A and B, respectively） were prepared, and the materials were incubated in cell culture fluid for 24 h.No material membrane orifice plate was used as blank control group.Contact angle meter was used to measure surface contact angle of protein membrane and composite membrane group（each group had three repeated holes）.Cell proliferation was assessed by cell counting kit-8 on the 4, 7, 14, and 21 days.The state of adhesion and growth of hDPSC on the materials surface was evaluated by scanning electron microscopy and cytoskeleton staining;and alkaline phosphatase （ALP） activity was measured to evaluate the cell differentiation potential.The expression of odontoblastic differentiation-related genes was measured by real-time PCR.Results Surface contact angle of the protein membrane group and composite membrane group A and group B were 89.51±0.12°, 70.32±0.07° and 71.31±0.09° respectively.hDPSC adhered well on each materials surface on the 7, 14, 21 days, ALP activity and differentiation genes of composite membrane group A and B rised more significantly than the blank control group and protein membrane group did （P〈0.05）.Dentin matrix protein1（DMP-1）, dentin sialoprotein（DSP）, ALP, osteoealcin（OC） mRNA expression reached peak on the 14 days in group A, and in group B on the 21 days.Bone sialoprotein（BSP） mRNA expression in both group A and B reached peak on the 21 days.Conclusions BG45S5-SF membrane is able to support the proliferation and showed the potential of odontoblastic differentiation for hDPSC.This finding suggests that BG45S5-SF membran

关 键 词：丝素蛋白 牙髓 干细胞 生物活性玻璃 

分 类 号：R783.1[医药卫生—口腔医学]